STAT proteins (signal transduction and activators of transcription) mediate a variety of extracellular signals. In response to ligand binding to receptor, these proteins become phosphorylated on an invariant tyrosine residue located near the C-terminus. Upon tyrosine phosphorylation, STAT proteins combine to form dimers, in which the SH2 domain of one monomer binds the pTyr of the second and the SH2 domain of the second monomer binds to the pTyr of the first. The dimer then migrates to the nucleus, binds to specific DNA promoter sequences and initiates transcription of designated response elements. In normal cells STATs mediate signaling from growth factor receptors and immune stimuli. However, in several cancer types, STATs are activated constitutively. In this proposal we will focus on Stat3, which has been shown to be constitutively activated in cancers of the breast, colon, and brain. The hypothesis of this proposal is that by blocking the dimerization of Stat3, we can eliminate the signaling mediated by this protein that promotes proliferation and prevents apoptosis and thereby inhibit the growth of tumor cells. To do this we intend to develop peptidomimetics based on phosphotyrosine containing peptides targeted to the Stat3 SH2 domain. We have demonstrated that a phosphopeptide derived from pTyr 705 of Stat3 is capable of inhibiting Stat3 activation in cells. In this grant we will develop inhibitors of Stat3 dimerization by 1. Optimizing amino acid sequences of Stat3 SH2 domain inhibitor peptides. 2. Put constrained tyrosines and other conformational constraints in smallest peptide to lock the conformation for optimal binding interactions. 3. Determine the structure of our peptides bound to the SH2 domain of Stat3 using X-ray crystallography and/or NMR to be used in refinement of our inhibitors. 4. Assay our compounds as inhibitors of dimerization and DNA binding, and test the most promising compounds in cell based assays of growth inhibition, soft agar colony formation, cell viability, and apoptosis.